The Retinal Determination pathway is a genetic cascade that was first described in the context of fly eye development. Much of what is known about this pathway comes from genetic and evolutionary studies. We have been involved in deciphering the molecular mechanisms by which the protein components of the RD pathway, Eyes Absent (EYA), Dachshund (DACH), and SIX, mediate their roles in cell-fate determination. The present proposal focuses on the Eyes Absent proteins, which we have shown to be protein tyrosine phosphatases in addition to their established transcriptional activation function. In flies, this activity contributes to eye development. However, the known phenotypes of Eya mutant mice suggest that EYAs may not play a comparable role in vertebrate eye development. In recent work we have correlated the tyrosine phosphatase activity of the EYAs with cell migration and invasion. Further EYAs are known to be expressed in cells of neural crest origin, and the anatomical defects reported for Eya1-/- mice are consistent with defects in neural crest migration. Thus our preliminary data suggests the novel hypothesis that the Eyes Absent proteins promote the cellular processes of motility and invasiveness that play a role in neural crest and eye development. This application is designed to decipher the signaling pathways that are regulated by the Eyes Absent proteins using in vitro cell culture methods as well as in vivo analysis of targeted Eya deletions in mice. An understanding of the mechanisms by which the EYAs function could potentially lead to new insights regarding their role in eye development, to an understanding of the potential role of EYAs in the development of eye diseases such as secondary cataracts and diabetic retinopathy, and to the development and spread of cancers. PUBLIC HEALTH RELEVANCE: The EYA proteins are part of a conserved regulatory pathway involved in embryonic eye development. Despite extensive genetic analyses in flies and mice the precise role of the EYA proteins in vertebrate eye development remains enigmatic. We have shown that the EYA proteins are both tyrosine phosphatases as well as transcriptional activators. Further we show that the phosphatase activity promotes cell migration and invasiveness. Based on this we propose that the EYAs play a role in the development of neural crest-derived ocular structures. In vitro and in vivo analyses of the molecular mechanisms by which the EYAs could affect both cell motility and the development of ocular structures of neural crest origin are proposed. The results from these studies could provide significant insight into the process of vertebrate eye development, as well as the mechanisms by which EYA mis-regulation could promote eye diseases and cancers.